Herpes viruses are responsible for significant disease both in immunocompetent and immunocompromised hosts. Despite the clinical importance of these viruses, little can be done to prevent herpes virus infections or reactivation of latent disease. To develop better strategies for prevention and treatment, a thorough understanding of the mechanism by which these viruses cause disease and by which certain cells and tissues are preferred targets for infection is critical. Recent studies, using fibroblast and epithelial cell lines, have shown that heparan sulfate (HS) glycosaminoglycans (GAGs) of cell surface proteoglycans serve as receptors for the human herpesviruses, herpes simplex virus type 1 (HSV-1) and herpes simplex virus type 2 (HSV-2). For HSV-1, it has been shown that the viral envelope glycoproteins, gC and gB, can independently mediate this attachment, although gC plays a key role. The viral glycoprotein(s) responsible for the binding of HSV-2 to the cell surface are not yet known. In addition, although HSV-1 and HSV-2 both bind to HS, it is not yet known whether the two serotypes of HSV bind to the same or different structural features of HS. Differences in receptors for the two serotypes may help explain differences in cell and tissue tropism, as well as differences in pathogenicity. The studies proposed in this grant application will define the specific structural features of the viral glycoproteins and cell surface GAGs important in the entry of HSV-1 and HSV-2 into cells. The aims of these studies are first to determine which glycoproteins mediate the binding of HSV-2 to cells by generating mutant viruses deficient in the potential heparin-binding glycoproteins. Second, the structural features of HS important for viral attachment of HSV-1 and HSV-2 will be defined using a series of chemically and enzymatically modified heparins. Compounds identified that differentiate binding of individual glycoproteins of both HSV-1 and HSV-2 to cell surface HS will be exploited in studies designed to map the heparin-binding domains of the individual glycoproteins. These studies will use a combination of recombinant viruses and site-directed mutagenesis. By biochemically and molecularly characterizing cell surface receptors and viral glycoproteins that mediate attachment of HSV- 1 and HSV-2 to cells, these studies should enhance our understanding of viral pathogenesis and tropism and may lead to the development of new approaches to antiviral therapy, vaccine development, and gene therapy.